V. Gopi

Detection of citrus yellow mosaic virus by PCR and nucleic acid spot hybridisation using non-radioactive probes in commercial citrus species

Citrus yellow mosaic virus (CYMV) was detected by polymerase chain reaction (PCR) in leaf samples of sweet orange, Rangpurlime, Pumello pink and acid lime and also in twig bark, fruit rind, fruit juice except fruit rag of sweet orange cv. Sathgudi, where all the positive samples showed bright amplification of the 726 bp band except in fruit rind. The CYMV could be detected by the biotin labelled probe of CYMV up to 1:100 dilutions by TE and TNE extraction methods in sweet orange cv. Sathgudi as well as Rangpurlime.

Symptom-based diagnosis of Huanglongbing (citrus greening) disease by PCR in sweet orange (Citrus sinensis Osbeck) and acid lime (Citrus aurantifolia Swingle)

Huanglongbing (HLB), previously known as citrus greening disease, is one of the most destructive diseases of citrus and responsible for the decline of citrus orchards in Andhra Pradesh (AP) and other citrus growing areas in the country. Polymerase chain reaction (PCR) amplification of 1160 bp fragment of 16S rDNA of HLB was observed in mottling symptoms, yellow vein symptoms, symptoms mixed with both citrus yellow mosaic virus and citrus greening and zinc deficiency-like symptoms; however, no amplification was observed in iron deficiency like symptoms. As the disease shows a variety of symptoms in infected trees, PCR detection could be useful for resolving confusion in identification of HLB between actual deficiency and deficiency like symptoms caused by HLB. In acid lime the midrib and veins of the leaves with both mottling and yellow vein symptoms were suitable for isolation of DNA and used as a template in the PCR test.

Finger printing of acid lime varieties and clones having varied resistance to bacterial canker, using RAPD marker

Citrus is an important fruit crop having divergent genetic variation within the species. The germplasm identification and characterisation is an important link between the conservation and utilisation of genetic resources. Conventionally, variety/clone identification has relied on morphological characters such as growth habit, leaf, floral and fruit characters etc. Investigation through RAPD (random amplified polymorphic DNA) markers was carried out for determination of genetic variation among 12 acid lime clones having varied resistance to bacterial canker disease. DNA was extracted from the leaf of 12 acid lime clones and was subjected to PCR using 20 random primers (nine from OPM and 11 from OPA series) which yielded a total of 127 distinct DNA fragments, out of which 103 were polymorphic. Genetic similarity was evaluated based on the presence or absence of bands. The bands obtained were polymorphic, with sizes ranging from 750 bp to 2.5 kb. Cluster analysis using the similarity coefficient showed that the Balaji, RHRL-124 and PKM-1 formed one cluster and the remaining clones formed a second cluster, which in turn were divided into TAL 94-9, TAL 94-10, TAL 94-11 and TAL 94-12 which formed the first subcluster; the Nalgonda selection and local acid lime formed a second subcluster; TAL 94-8, RHRL-49 and RHRL-122 did not resemble any other clones. Among the 12 acid lime clones, Balaji, RHRL-124, RHRL-122 and PKM-1 were found to be moderately resistant to bacterial canker. Correlation of RAPD data with canker disease incidence in the moderately resistant acid lime clones viz., Balaji, RHRL-124 and PKM-1 were formed as one cluster, and all susceptible clones formed as a second cluster viz., except TAL-94-9, RHRL-122, which were found to be moderately resistant and did not form a cluster with any other acid lime clone.

Variability in Protein and Esterase Profiles of Botryodiplodia theobromae an Incitant of Longitudinal Splitting of Bark and Wood Disease of Acidlime (Citrus aurantifolia Swingle)

Acid lime (Citrus aurantifolia Swingle) is one of the important citrus fruit grown over an area of 0.90m ha and constitutes nearly 20% of the total citrus production 90.75mts in India. In India, Andhra Pradesh, Maharashtra, Karnataka, Punjab, and Assam are the leading acid lime growing states. Among the states, A.P occupies the first place both in area and production of limes followed by Maharashtra.

Improvement of antagonism and carbendazim tolerance in native Trichoderma isolates through ethyl methane sulfonate (EMS) mutagenesis

Genetic enhancement of TCT4 and TCT10 was aimed in the present paper. Trichoderma reesei (TCT10/M18) mutant isolate evolved by ethyl methane sulfonate mutations was found to exhibit altered properties compared to its parent isolates. This mutant grew well in the potato dextrose agar (PDA) medium containing carbendazim (50 ppm). RAPD-PCR results suggested the uniqueness of mutants, which was useful in differentiating mutant and wild Trichoderma isolates. These mutants established well in the rhizosphere of rough lemon seedlings. The seedlings treated with carbendazim followed by an application of carbendazim-resistant mutant (TCT10/M18) resulted in a better seedling emergence and a less dry root rot disease caused by Fusarium solani in nursery conditions.